Team:Czech Republic/Goals
Goals
Module 1 builds synthetic haploid strains with refactored mating loci that are conjugated to make a functional IOD. These strains have the wild-type mating phenotype and differentially express a reprogrammed signalling pathway in their diploid state proving the feasibility of the clone-free assembly concept.
- ✓ Construct a set of reporter promoters for yeast cells
- ✓ Characterize reporter promoters in all mating types
- ✓ Design and materialized synthetic MATa and MATx strains
- ✓ Build a synthetic diploid strain with a functional yeast pheromone pathway
- ✓ Demonstrate the correct functionality of yeast pheromone pathway in synthetic diploids
Module 2 builds a set of orthogonal pheromones and receptors. These pheromone-receptor pairs enable specific localized signalling proving the feasibility of multichannel signal transmission underlying logic operations necessary for reliable diagnosis.
- ✓ Construct a set of yeast plasmids with different mating pheromones and their receptors and contribute to the Registry with BioBricks
- ✓ Verify the correct coupling of the receptors to the yeast pheromone mating pathway
- ✓ Verify the correct expression and secretion of the different pheromones
- ✓ Show the orthogonality of the used receptors and pheromones
Module 3 builds a set of location tags that recognize common tumor surface markers and agglutinate cell populations. Location tags displayed in the correct conformation strengthen cell-cell interactions to enable localization of signal transmission.
- ✓ Express streptavidin, EpCAM, Anti-EpCAM scFv, c-Myc scFv and anti-HuA scFv on the surface of yeasts
- ✓ Monitor the dynamic binding of the displayed fragments of antibodies and corresponding markers
- ✓ Demonstrate compatibility of our yeast system and blood and functionality of the location tags in plasma
Module 4 provides modeling and simulation support for other modules.
- ✓ Develop a simulation environment CeCe to capture the complexity of cell-cell signal transmission
- ✓ Design an IOD chemical reaction network model
- ✓ Develop a schematic architecture for conceptual modeling of signal transmission networks
- ✓ Design a two IOD signal transmission network suitable for the IOD band
- ✓ Illustrate the robustness and efficiency of the IOD band design in CeCe simulations
Module 5 allows other modules to use microfluidics devices for their experiments.
- ✓ Set of microfluidic devices fabricated by PDMS soft-lithography.
- ✓ Characterization of signal transmission range between wildtype MATa and MATx Saccharomyces cerevisiae cells.
- ✓ Dynamic characterisation of signal transmission between synthetic MATa and MATx Saccharomyces cerevisiae cells.
- ✓ Comparison with mathematical model of signal transmission mechanism and estimation of the activation threshold for different cell concentrations.
- ✓ Demonstrate yeast induced blood agglutination on-chip by human antigen A displayed on cell surface by Yeast Surface Display.